Liquid chromatography is a well-known procedure used in analytical and preparative chemistry. It is most commonly carried out in an elongated cylinder, usually formed of stainless steel, in which a porous chromatographic packing is immobilized. The sample to be analyzed is fed into the column in a liquid carrier. As the sample passes through the column, various components thereof are delayed for differing times because of differing interaction (chemical or physical) with the packing material. The effluent is continuously monitored to identify components as they emerge from the column.
The reproducibility of the process depends in large part on the column packing remaining in a constant condition. In the past, the primary emphasis on achieving the most dependable packing beds has been on the use of improved processes for putting the packing in place.
Many techniques have been suggested including vibration (See U.S. Pat. No. 3,300,849): All of these techniques require careful control if segregation of particles by size is to be avoided and uniformly packed columns are to be obtained. In general, the most commonly used practice of filling a high-performance column has been a costly method including slurrying the packing and passing the slurry into the column; thereby, in effect, using the column itself as a form for placing a "filter cake" of chromatographic packing therein.
A number of solutions have been suggested for holding the packing "in-place". Some of these, like the aforementioned vibration technique and slurrying technique, emphasize a maximum effort to put a conventional packing into the column in such a way as to have it assume a stable position. Other techniques, such as those described in U.S. Pat. No. 3,808,125 to Good, are rather complex or expensive procedures for fastening the packing to the column wall.
None of these attempts by the prior art have been dependably successful in achieving any of an excellent performance, a column-to-column consistency in separating characteristics, or a desired degree of stability of performance over a period of time for a single column at a cost which can make the apparatus available to the broadest spectrum of chromatographers.
In discussing packed-column processes, it is helpful to recognize four kinds of space, all of which can be referred to as "void volume." These include (1) void volume inside a porous particle; (2) theoretical void volume between particles, i.e. the type of unavoidable volume which would result from a perfectly efficient packing of spheres of the same size; (3) void volume which is attributable to imperfect packing of particles, usually present to some extent in any actual system utilizing a particulate-packing system; and (4) void volume which represents relatively large voids resulting from the consolidation of those voids described in (3). Void volume as generally used herein relates to a composite of void volumes (3) and (4).
It should be realized that the description of the prior art which is set forth above is, necessarily, made in hindsight and in view of knowledge contributed only by the instant invention. Nothing in the description is intended to be construed as an indication that the state of the prior art as described above was, or could have been, appreciated by those of ordinary skill in the art before their knowledge of the invention described below.